The proposed work will develop the hypothesis that the concentration of acetylcholine (ACh) at postjunctional effector sites in the atrium regulates the expression of beta-adrenergic-cyclic AMP mediated functions. Isoproterenol stimulated cyclic AMP accumulation in isolated murine atria is markedly inhibited by K ion-induced depolarization, cholinesterase inhibition or carbachol. All of these manipulations block cyclic AMP accumulation through a muscarinic (atropine-sensitive) mechanism. I propose to demonstrate that elevated K ion releases ACh from prelabeled (with 3H-choline) neurotransmitter stores in isolated atria, and that subsequent inhibition of cyclic AMP accumulation is proportional to the available released ACh. By quantitation of the effect of released or exogenous ACh, I will determine whether there is noncompetitive antagonism of catecholamine stimulated cyclic AMP accumulation by muscarinic receptor activation. Isoproterenol induced activation of protein kinase and phosphorylase will also be shown to be inhibited in proportion to the concentration of choline ester. I will examine local presynaptic mechanisms by which drugs and hormones might modulate K ion-induced ACh release, as potential physiological regulators of the expression of cyclic AMP dependent beta-adrenergic functions. Developmental and hormonal changes in the density or sensitivity of muscarinic receptors will be utilized to examine post-synaptic cholinergic receptor mechanisms regulating adrenergic responses. Other studies will focus on regulation of muscarinic receptors and will ask whether separate populations of muscarinic receptors regulate cyclic AMP formation, phospholipid metabolism, catecholamine release and atrial rate. The eventual goals of this work will be to demonstrate that some parasympathetic effects in the atrium can be explained by cholinergic inhibition of cyclic AMP accumulation, while others may result from change in membrane phospholipid metabolism, and to elucidate the role of intracardiac cholinergic neurons as critical effector sites for drug and hormone actions on cardiac muscle.